The long term goal of my research is to understand the mechanisms of K about channels, a group of highly diversified integral membrane proteins. A special class of K+ channels, called inward about rectifiers, pass K+ ions in the inward direction more efficiently than in the outward direction. This property, referred to as inward rectification, allows inward-rectifier K+ channels to maintain and regulate the resting membrane potential and thereby accomplish many important biological tasks. The experiments proposed here are primarily aimed to understand the mechanisms underlying inward rectification, i.e., how the channels are blocked by intracellular cations. The channels will be expressed in Xenopus oocytes and their functional properties will be examined using electrophysiological techniques in conjunction with mutagenesis. Aim 1: To understand the mechanisms of inward rectifier block by intracellular cations we will determine: a) the kinetic mechanism of channel block by intracellular polyamines, b) the mechanism which confers high affinity to the interaction of inward rectifiers with intracellular polyamines, and c) the molecular mechanism underlying the channel's specificity for intracellular cations. In Aim 2, we will use the conduction pore of a bacterial K+ channel (KcsA)-whose structure has been determined with X-ray diffraction-as a model to further examine the mechanism by which the channel achieves the high-affinity binding of intracellular cations. The proposed studies have important medical implications. Inward-rectifier K+ channels play many important biological roles, e.g., they mediate the control of heart rate by the vagus nerve, modulate synaptic transmission in the nervous system, and couple blood glucose levels to insulin secretion. Thus, these channels represent important pharmacological targets for medical intervention during various disease states.